Novel carbonyl and nitrile products from reactive chlorinating species attack of lysosphingolipid

Lysosphingolipids are important lipid signaling molecules that are associated predominantly with high density lipoproteins (HDL) in human plasma. Further, HDL has been shown to be a target for the reactive chlorinating species (RCS) produced by myeloperoxidase (MPO). Accordingly, RCS attack of lysosphingolipids was characterized in these studies. It was shown that RCS attack of sphingosylphosphorylcholine results in the formation of 2-hexadecenal and 1-cyano methano phosphocholine. The structures were identified and confirmed predominantly using mass spectrometric analyses. Further, it was demonstrated that RCS attack of another bioactive lysosphingolipid sphingosine 1-phosphate also results in the formation of 2-hexadecenal from its sphingosine base. Using a synthetically prepared, deuterated 2-hexadecenal internal standard, it was determined that 2-hexadecenal quickly accumulated in HDL treated with MPO/RCS generating system. Thus, the present studies characterize the formation of a novel group of lipid products generated following RCS attack of lysosphingolipids.     

Agonistic monoclonal antibody against CD40 receptor decreases lymphocyte apoptosis and improves survival in sepsis

Sepsis causes a marked apoptosis-induced depletion of lymphocytes. The degree of lymphocyte apoptosis during sepsis strongly correlates with survival. CD40, a member of the TNFR family, is expressed on APCs and has potent antiapoptotic activity. In this study we determined whether an agonistic Ab against CD40 could protect lymphocytes from sepsis-induced apoptosis. Secondly, we examined potential antiapoptotic mechanisms of the putative protection. Lastly, we aimed to determine whether anti-CD40 treatment could improve survival in sepsis. CD1 mice were made septic by the cecal ligation and puncture method and treated postoperatively with anti-CD40 Ab. Treatment with anti-CD40 completely abrogated sepsis-induced splenic B cell death and, surprisingly, decreased splenic and thymic T cell death as well (p < 0.001). To investigate the mechanism of protection of anti-CD40 therapy on T cells, CD40 receptor expression was examined. As anticipated, the CD40 receptor was constitutively expressed on B cells, but, unexpectedly, splenic and thymic T cells were found to express CD40 receptor during sepsis. Furthermore, CD4+CD8- T cells were the predominant subtype of T cells expressing CD40 receptor during sepsis. Additionally, the antiapoptotic protein Bcl-x(L) was found to be markedly increased in splenic B and T cells as well as in thymic T cells after treatment with anti-CD40 Ab (p < 0.0025). Lastly, mice that were made septic in a double injury model of sepsis had improved survival after treatment with anti-CD40 as compared with controls (p = 0.05). In conclusion, anti-CD40 treatment increases Bcl-x(L), provides nearly complete protection against sepsis-induced lymphocyte apoptosis, and improves survival in sepsis.     

Complete physical map of the rfb gene cluster encoding biosynthetic enzymes for the O antigen of Salmonella typhimurium LT2.

Biosynthesis of the Salmonella typhimurium LT2 O antigen is encoded by genes which map in the rfb cluster. The cloning and restriction enzyme analysis of part of this cluster have been described previously (H. N. Brahmbhatt, N. B. Quigley, and P. R. Reeves, Mol. Gen. Genet. 203:172-176, 1986). The entire rfb gene cluster has now been cloned, and a detailed restriction enzyme map has been constructed which has enabled us to map the approximate positions of individual rfb genes.     

Cloning part of the region encoding biosynthetic enzymes for surface antigen (O-antigen) of Salmonella typhimurium

Summary The rfb gene cluster of Salmonella typhimurium encodes the enzymes required for the biosynthesis of the O-Antigen. A part of it has been cloned in plasmid vectors pBR322 and pUC9 using an adjacent, previously cloned, part of the his operon (Barnes 1981) as a molecular probe for the first clone. A detailed restriction enzyme map of 7.57 kb of rfb DNA is presented and the approximate locations of two of the genes, rfbK and rfbM have been defined.     

Microwave-assisted solvent-free synthesis of some novel thiazole-substituted thiosemicarbazone analogues: antimicrobial and anticancer studies

The increased resistance to antibiotics has compelled researchers to devise novel active compounds targeting multidrug-resistant pathogenic microorganisms. A series of thiosemicarbazone derivatives was synthesized by reacting thiosemicarbazide with 2-aryl-4-formylthiazole, 2-aryl-5-formyl-4-methylthiazole, and/or 5-acetyl-2-aryl-4-methylthiazole compounds. These thiosemicarbazone-based thiazole adducts were evaluated for their inhibitory activities against tuberculosis H37Ra and Bovis BCG mycobacteria. Their cytotoxicity was assessed against two cancer cell lines: colonic carcinoma (HCT-116) and cervical cancer (HeLa). Notably, these thiosemicarbazones exhibited minimal cytotoxic effects on these cell lines even at their highest concentrations. Furthermore, the prepared thiosemicarbazone derivatives demonstrated significant antimicrobial efficacy against Bacillus subtilis and Staphylococcus aureus (Gram-positive bacterial pathogens) as well as Escherichia coli and Pseudomonas fluorescens (Gram-negative bacterial pathogens). While most of the prepared thiosemicarbazone derivatives exhibited moderate activity against Candida albicans (a fungal strain), their performance was notable. The thiosemicarbazone-based thiazole adducts were also successfully synthesized using a solvent-free approach under microwave irradiation. Compared with conventional reflux methods, the microwave-assisted technique yielded high thiazole yields within just 5 min, obviating the need for catalysis. This study signifies significant strides toward the rational design of more potent antimycobacterial agents.     

Associations between women’s empowerment and child development,growth, and nurturing care practices in sub-Saharan Africa: A cross-sectional analysis of demographic and health survey data

BackgroundApproximately 40% of children 3 to 4 years of age in low- and middle-income countries have suboptimal development and growth. Women’s empowerment may help provide inputs of nurturing care for early development and growth by building caregiver capacity and family support. We examined the associations between women’s empowerment and child development, growth, early learning, and nutrition in sub-Saharan Africa (SSA).Methods and findingsWe pooled data on married women (15 to 49 years) and their children (36 to 59 months) from Demographic and Health Surveys that collected data on child development (2011 to 2018) in 9 SSA countries (N = 21,434): Benin, Burundi, Cameroon, Chad, Congo, Rwanda, Senegal, Togo, and Uganda. We constructed a women’s empowerment score using factor analysis and assigned women to country-specific quintile categories. The child outcomes included cognitive, socioemotional, literacy–numeracy, and physical development (Early Childhood Development Index), linear growth (height-for-age Z-score (HAZ) and stunting (HAZ <−2). Early learning outcomes were number of parental stimulation activities (range 0 to 6) and learning resources (range 0 to 4). The nutrition outcome was child dietary diversity score (DDS, range 0 to 7). We assessed the relationship between women’s empowerment and child development, growth, early learning, and nutrition using multivariate generalized linear models.On average, households in our sample were large (8.5 ± 5.7 members) and primarily living in rural areas (71%). Women were 31 ± 6.6 years on average, 54% had no education, and 31% had completed primary education. Children were 47 ± 7 months old and 49% were female. About 23% of children had suboptimal cognitive development, 31% had suboptimal socioemotional development, and 90% had suboptimal literacy–numeracy development. Only 9% of children had suboptimal physical development, but 35% were stunted. Approximately 14% of mothers and 3% of fathers provided ≥4 stimulation activities. Relative to the lowest quintile category, children of women in the highest empowerment quintile category were less likely to have suboptimal cognitive development (relative risk (RR) 0.89; 95% confidence interval (CI) 0.80, 0.99), had higher HAZ (mean difference (MD) 0.09; 95% CI 0.02, 0.16), lower risk of stunting (RR 0.93; 95% CI 0.87, 1.00), higher DDS (MD 0.17; 95% CI 0.06, 0.29), had 0.07 (95% CI 0.01, 0.13) additional learning resources, and received 0.16 (95% CI 0.06, 0.25) additional stimulation activities from their mothers and 0.23 (95% CI 0.17 to 0.29) additional activities from their fathers. We found no evidence that women’s empowerment was associated with socioemotional, literacy–numeracy, or physical development. Study limitations include the possibility of reverse causality and suboptimal assessments of the outcomes and exposure.ConclusionsWomen’s empowerment was positively associated with early child cognitive development, child growth, early learning, and nutrition outcomes in SSA. Efforts to improve child development and growth should consider women’s empowerment as a potential strategy.

Lilia Bliznashka and co-workers study empowerment of women and associated child health outcomes in sub-Saharan Africa.     

Leukemia inhibitory factor: Role in human mesenchymal stem cells mediated immunosuppression

The interactions between mesenchymal stem cells (MSCs) and immune system are currently being explored. Leukemia inhibitory factor (LIF) is linked to regulatory transplantation tolerance. Our aim was to study the expression of LIF on human MSCs at both gene and protein level in mixed lymphocyte reaction (MSC/MLR), and its implication in MSC immunosuppressive effect. There was a 7-fold increase (611pg/ml) in LIF in MSC/MLR as compared to MSCs alone. Using LIF neutralizing antibody, a significant restoration of up to 91% of CD3+ lymphocyte proliferation in MSC/MLR was observed (p=0.021). LIF was implicated in the generation of regulatory lymphocytes, as demonstrated by decrease of Foxp3+ regulatory cells after using LIF neutralizing antibody in MSC/MLR (p=0.06) by flow cytometry. A positive correlation between LIF and human leukocyte antigen (HLA-G) gene expression by MSCs was found (R(2)=0.74). Our findings provide evidence supporting the immunomodulatory effect of MSCs.     

Expression of the gene for mouse lactate dehydrogenase C (Ldhc) is required for male fertility

The lactate dehydrogenase (LDH) protein family members characteristically are distributed in tissue- and cell type-specific patterns and serve as the terminal enzyme of glycolysis, catalyzing reversible oxidation reduction between pyruvate and lactate. They are present as tetramers, and one family member, LDHC, is abundant in spermatocytes, spermatids, and sperm, but also is found in modest amounts in oocytes. We disrupted the Ldhc gene to determine whether LDHC is required for spermatogenesis, oogenesis, and/or sperm and egg function. The targeted disruption of Ldhc severely impaired fertility in male Ldhc(-/-) mice but not in female Ldhc(-/-) mice. Testis and sperm morphology and sperm production appeared to be normal. However, total LDH enzymatic activity was considerably lower in Ldhc(-/-) sperm than in wild type sperm, indicating that the LDHC homotetramer (LDH-C(4)) is responsible for most of the LDH activity in sperm. Although initially motile when isolated, there was a more rapid reduction in the level of ATP and in motility in Ldhc(-)(/-) sperm than in wild-type sperm. Moreover, Ldhc(-/-) sperm did not acquire hyperactivated motility, were unable to penetrate the zona pellucida in vitro, and failed to undergo the phosphorylation events characteristic of capacitation. These studies showed that LDHC plays an essential role in maintenance of the processes of glycolysis and ATP production in the flagellum that are required for male fertility and sperm function.     

Design of an engineered N-terminal HIV-1 gp41 trimer with enhanced stability and potency

HIV fusion is mediated by a conformational transition in which the C-terminal region (HR2) of gp41 interacts with the N-terminal region (HR1) to form a six-helix bundle. Peptides derived from the HR1 form a well-characterized, trimeric coiled-coil bundle in the presence of HR2 peptides, but there is little structural information on the isolated HR1 trimer. Using protein design, we have designed synthetic HR1 peptides that form soluble, thermostable HR1 trimers. In vitro binding of HR2 peptides to the engineered trimer suggests that the design strategy has not significantly impacted the ability to form the six-helix bundle. The peptides have enhanced antiviral activity compared to wild type, with up to 30-fold greater potency against certain viral isolates. In vitro passaging was used to generate HR1-resistant virus and the observed resistance mutations map to the HR2 region of gp41, demonstrating that the peptides block the fusion process by binding to the viral HR2 domain. Interestingly, the activity of the HR2 fusion inhibitor, enfuvirtide (ENF), against these resistant viruses is maintained or improved up to fivefold. The 1.5 A crystal structure of one of these designs has been determined, and we show that the isolated HR1 is very similar to the conformation of the HR1 in the six-helix bundle. These results provide an initial model of the pre-fusogenic state, are attractive starting points for identifying novel fusion inhibitors, and offer new opportunities for developing HIV therapeutics based on HR1 peptides.